URINARY CRYSTAL DETECTION METHOD

Abstract

The present invention provides an urinary crystal detection method, which is used with a crystal collecting unit and a crystal detecting unit. The method comprises steps of using magnetic particles to attach urinary crystals from a sample. Then, providing a magnetic field to separate the urinary crystals. Further to analyz their constituent by the Raman signals of the urinary crystal to access an urinary calculus result for urinary stone patient.

Claims

1. An urinary crystal detection method, comprising steps: providing a urine sample from a patient; adding a plurality of magnetic particles into the urine sample, the magnetic particles attaching to a plurality of urine crystals of the urine sample, wherein the magnetic particles are formed as a core, which is coated with gelatin, and the surfaces of the magnetic particles are modified by PO(OH).sub.2 or PO.sub.4.sup.3− groups to attach the urinary crystals with Ca or Mg; providing a magnetic field to attract the magnetic particles by a magnetic separating device, and collecting the urinary crystals which are absorbed by the magnetic particles; analyzing a Raman signal of the urinary crystals by a microscopic Raman spectroscopy unit; comparing the peak characteristics of the Raman signal with a component information from a database, wherein the component information is a plurality of reference peak characteristics from different urinary crystals related to a plurality of urinary calculus; and receiving a result of the urinary calculus which the reference peak characteristics is similar as the Raman signal.

2. The urinary crystal detection method of claim 1, wherein the component of the magnetic particles are formed by Fe.sub.3O.sub.4 coated with gelatin or PSMA.

3. The urinary crystal detection method of claim 1, wherein the size of the magnetic particle is 5 nm˜200 nm.

4. The urinary crystal detection method of claim 1, wherein the gelatin are modified by FITC.

5. The urinary crystal detection method of claim 1, wherein the magnetic separating device is a magnet or a magnetite.

6. The urinary crystal detection method of claim 1, wherein the steps further comprise processing the urine sample before the step of adding the magnetic particles into the urine sample, includes: detecting the urine sample to get a pH value ; and executing processes according to the pH value, if the pH value of urine sample is less than 7, the urine sample is executed an acid processes; if the pH value of urine sample 10 is larger than 7, the urine sample is executed an alkali processes.

7. The urinary crystal detection method of claim 8, wherein the acid processes comprise a step of adding 5%˜15% acetic acid to the urine sample.

8. The urinary crystal detection method of claim 8, wherein the alkali processes comprise a step of adding 5%˜15% NaOH to the urine sample.

9. The urinary crystal detection method of claim 1, wherein the steps further comprises analyzing an auto-fluorescence signal of the urinary crystals by a fluorescence spectroscopy unit.

10. The urinary crystal detection method of claim 1, wherein the surfaces of the magnetic particles are modified by a plurality of fluorescent molecules.

11. The urinary crystal detection method of claim 10, wherein the fluorescent molecules are PAR or CV.

12. The urinary crystal detection method of claim 11, wherein the steps further comprises analyzing a fluorescence signal of the fluorescent molecules by a fluorescence spectroscopy unit.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0034] FIG. 1 is a diagram of the urinary crystal detection system in the invention.

[0035] FIG. 2 is an embodiment diagram of the urinary crystal detection system in the invention.

[0036] FIG. 3A is a diagram of the magnetic particles in the invention.

[0037] FIG. 3B shows an embodiment diagram of the magnetic particles in the invention.

[0038] FIG. 3C shows a diagram of a third embodiment of the magnetic particle in the invention.

[0039] FIG. 3D shows a diagram of a fourth embodiment of the magnetic particle in the invention.

[0040] FIG. 4A shows an electron microscope diagram of the magnetic particles with the size of about 200 nm.

[0041] FIG. 4B shows an electron microscope diagram of the magnetic particles with the size of about 13 nm.

[0042] FIG. 5A is a diagram showing pure substance powder of Calcium oxalate.

[0043] FIG. 5B is a diagram showing the magnetic particles formed by Fe.sub.3O.sub.4 added surface-modified. PO.sub.4.sup.3−. The magnetic particles are adsorbed onto the Calcium oxalate in urine. FIG. 5C is a diagram showing the magnetic particles and the adsorbed Calcium oxalate attracted by an external magnet.

[0044] FIG. 6 shows a diagram of the magnetic particles adsorbed onto Calcium oxalate.

[0045] FIG. 7 shows a flow chart of the urinary crystal detection method.

[0046] FIG. 8A is a diagram of urinary crystals under the microscope, and the status of the urinary crystals is calcium oxalate dihydrate (COD). FIG. 8B is a diagram of Raman signals of the COD.

[0047] FIG. 9A is a diagram of urinary crystals under the microscope, and the status of the urinary crystals is hydroxyapatite (HAP).

[0048] FIG. 9B is a diagram of Raman signals of the HAP.

DETAILED DESCRIPTION OF THE INVENTION

[0049] For clarity of disclosure, and not by way of limitation, the detailed description of the invention is divided into the subsections that follow.

[0050] The present invention provides a method, which is used with a urinary crystal detection system, can collect and detect urinary crystals from patients' urine samples, so that different patients can have different treatments according to the types of the urinary calculus, which can be further determined from the type of the urinary crystal.

[0051] Please refer to FIG. 1 showing a diagram of the urinary crystal detection system in the invention. The urinary crystal detection system includes a crystal collecting unit 30 and a crystal detecting unit 40. Also, the crystal collecting unit 30 includes a magnetic separating device 310.

[0052] An urine sample 10 of a patient is received by the crystal collecting unit 30, and is added magnetic particles. The magnetic separating device 310 attracts the magnetic particles attached to the urinary crystals, and disposes the magnetic particles and transfers the urinary crystals to a carrier, such as Slides, quartz slides and so on. The crystal detecting unit 40 detects the type of the urinary crystal.

[0053] The crystal detecting unit 40 can be a microscopy, a Raman spectroscopy, an infrared spectroscopy, a fluorescence spectroscopy, a X-ray diffraction (XRD) or image analyzer, which is not limited herein. In a preferred embodiment, the crystal detecting unit 40 is a Raman spectroscopy to measure Raman signals of the urinary crystals and further to determine the type of the urinary crystals.

[0054] The urinary crystal detection system also comprises a sample processing unit 20. The sample processing unit 20 including a pH detecting device 210 and pH processing device 220. The pH detecting device 210 measures the pH value of the urine sample 10. The urine sample 10 is detected by the pH detecting device 210 to generate a pH value, and executed processes by the pH processing device 220 according to the pH value. If the pH value of urine sample 10 is less than 7, the urine sample 10 is executed the acid processes; if the pH value of urine sample 10 is larger than 7, the urine sample 10 is executed the alkali processes. The acid processes and the alkali processes are used to separate the proteins, cells and etc. from the urinary crystals.

[0055] The urinary crystal detection system of the present invention can include or exclude the sample processing unit 20. The urine sample 10 can be directly used in the urinary crystal detection system of the present invention, or executed a pre-treatment by the sample processing unit 20 before the crystal collecting unit 30 receiving the urine sample 10. Moreover, a user can directly provide a pre-treated urine sample 10 for the urinary crystal detection system of the present invention, which is not limited herein.

[0056] Please refer to FIG. 2 showing a diagram of another embodiment of the urinary crystal detection system in the invention. The urine sample 10 is detected by the pH detecting device 210 to generate a pH value, and executed processes according to the pH value. The amount of the urine sample 10 is 1-5 ml from a patient's urine, which is not limited herein.

[0057] In an embodiment, when the pH value of the urine sample 10 detected is larger than a threshold, the urine sample 10 is added alkali solution. For example, the pH value of the threshold pH=7, the pH value from urine sample 10 detected is larger than 7, the urine sample 10 is added 5%˜15% NaOH. In a preferred embodiment, the weight percentage concentration of NaOH is 10%, and the volume of NaOH is 200 μl, which is not limited herein.

[0058] In an embodiment, when the pH value from the urine sample 10 is lower than a threshold, the urine sample 10 is added acid solution. For example, the pH value of the threshold is 7, the pH value from the urine sample 10 is lower than 7, the urine sample 10 is added 5%˜15% acetic acid. In a preferred embodiment, the weight percentage concentration of acetic acid is 10%, and the volume of acetic acid is 200 μl, which is not limited herein.

[0059] The criteria of the pH value of the threshold can be adjusted by users. The value of pH=7 is an example in the invention, but not limited herein.

[0060] In an embodiment, after the urine sample 10 is executed above-mentioned acid or alkali processes, the urine sample 10 containing urinary crystals is added buffer solution, such as adding 5 ml of secondary water or three-times distilled water, to clean and remove impurities, which is not limited herein.

[0061] Then, the urinary crystals from urine sample 10 are received by the crystal collecting unit 30. In an embodiment, a plurality of magnetic particles are added to the urine sample 10, and the magnetic particles adsorb onto urinary crystals. The magnetic separating device 310 attracts the magnetic particles to collect the urinary crystals adsorbed by the magnetic particles. The collected magnetic particles and the urinary crystals are disposed on a carrier, such as slides, quartz slides and so on.

[0062] The magnetic separating device 310 disposes the urinary crystals on the carrier, such as quartz slides. The carrier is received by the crystal detecting unit 40 to execute detecting processes. The crystal detecting unit 40 can be any detector, such as a microscopy, a Raman spectroscopy, an infrared spectroscopy, a X-ray diffraction (XRD) or image analyzer, which is not limited herein. In a preferred embodiment, the crystal detecting unit 40 is a Raman spectroscopy to measure Raman signals from urinary crystals, which is not limited herein.

[0063] The crystal detecting unit 40 comprise a database 410. The database 410 is used to store the component information of all kinds of urinary crystal and urinary calculus. The urinary crystal is detected by the crystal detecting unit 40 to generate a detecting result. The detecting result is compared with the database 410 to determine the type of the urinary crystal. In a preferred embodiment, the crystal detecting unit 40 is a Raman spectroscopy to measure the Raman signals of the urinary crystals. The peak characteristics of Raman signals are compared with the component information of the database 410 and further to determine the type of the urinary crystals, which is not limited herein.

[0064] In an embodiment, the magnetic particles 50 are formed by a plurality of core 510 coated with protective layers 520. Please refer to FIG. 3A showing a diagram of the magnetic particle in the invention. The components of the cores 510 can be Fe, Co, Ni , FeO.sub.2, Fe.sub.2O.sub.3 or Fe.sub.3O.sub.4, which is not limited herein. The components of the protective layers 520 can be a surfactant, a polymer layer, a carbon/silicon inorganic layer, collagen or gelatin. In a preferred embodiment, the component of the cores 510 of the magnetic particles 50 is Fe.sub.3O.sub.4 coated with gelatin as the protective layer 520. In a preferred embodiment, the component of the cores 510 of the magnetic particles 50 is Fe.sub.3O.sub.4 coated with PSMA as the protective layer 520, which is not limited herein.

[0065] The synthesis methods of the magnetic particles including coprecipitation method, thermal-chemical-vapor-deposition method, reduction method, micellar method, hydrothermal method and so on. Various synthetic methods are studied by scholars, which is not limited herein.

[0066] The main components of urinary calculi are calcium, magnesium, uric acid (such as calcium oxalate, magnesium ammonium phosphate). In a preferred embodiment, the magnetic particles 50 is Fe.sub.3O.sub.4 coated with gelatin (containing a benzene ring), and the component of the gelatin is animal collagen, which is a kind of natural high molecular polypeptide polymer. Therefore, the protective layer 520 composed of gelatin as magnetic particles 50 can absorb most kinds of urinary crystals. In a situation of adsorbing a specific type of urinary crystal, the surface of the magnetic particles 50 can be further modified a functional group to make the magnetic particles 50 can select types of urinary crystal, such as shown in FIG. 3B.

[0067] In another embodiment of the invention, the component of the cores 510 of the magnetic particles 50 is Fe.sub.3O.sub.4 coated with PSMA or gelatin as the protective layer 520. The PSMA includes benzene ring structure to adsorb most kinds of organic urinary crystals, such as adsorbing uric acid crystals, but which is not limited herein.

[0068] In an embodiment of the invention, the magnetic particles 50 are formed by Fe.sub.3O.sub.4 and coated with surface-modified PO.sub.4.sup.3−, as shown in FIG. 3B. The characteristic of R—PO.sub.4.sup.3− molecule is easy to bind with calcium ions or magnesium ions, thus the functional group of R—PO.sub.4.sup.3− have chemical bonding with the surfaces of magnetic nanoparticles of Fe.sub.3O.sub.4 . The surfaces of nanoparticles with functional PO.sub.4.sup.3− group can further to form a covalent bond with calcium and magnesium. That is, the molecular structure is easy to have chemical reaction and bind with urine acid molecules, and it is effective to separate and purify urinary crystals with calcium or magnesium. Another example of magnetic particles 50 coated with the gelatin which structure contain benzene ring-rich functional group and hydrogen bond can be used for the adsorption of organic urinary crystals, such as adsorption of uric acid crystals, which is not limited herein.

[0069] In another embodiment, the surfaces of the magnetic particles 50 are modified by fluorescent molecules 540, such as Pararosaniline hydrochloride (PAR), Crystal Violet (CV) or Fluorescein isothiocyanate (FITC), as shown in FIG. 3C. After adsorbing urinary crystals, the magnetic particles 50 modified by fluorescent molecules 540 can mark the uric acid crystals. It is beneficial to quickly and accurately detect the types of the urinary crystals and it also can be used for automated detecting apparatuses. The automated detecting apparatuses can detect the urinary crystals via the fluorescent molecules. Any component having a benzene ring structure, NH.sub.2 functional group or fluorescent molecules with positive charge can be used, the present invention is not limited thereto.

[0070] Furthermore, in an embodiment, in the FIG. 3D, the surfaces of the magnetic particles 50 can be modified by PO.sub.4.sup.3− 530 and fluorescent molecules 540 at the same time to make the magnetic particles 50 can have the functions of determining types of the urinary crystals and marking the urinary crystals.

[0071] The size of magnetic particles is 5-200 nm, Please refers to FIG. 4A and FIG. 4B showing the electron microscope diagrams of different sizes of the magnetic particles. The size of magnetic particles in FIG. 4A is about 200 nm and the size of magnetic particles in FIG. 4B is about 13 nm. The preferred size of magnetic particle is 13 nm, which is not limited herein.

[0072] In above embodiment, please refer to FIG. 5A to FIG. 5C showing diagrams of surface-modified PO.sub.4.sup.3− surface-modified of Fe.sub.3O.sub.4 magnetic particles used to adsorb calcium oxalate. FIG. 5A is a diagram showing pure substance powder of Calcium oxalate. FIG. 5B is a diagram showing the magnetic particles formed by Fe.sub.3O.sub.4 added in surface-modified PO.sub.4.sup.3− groups. The magnetic particles adsorb onto the Calcium oxalate. FIG. 5C is a diagram showing the magnetic particles and the Calcium oxalate attracted by an external magnet. Please refer to FIG. 6 showing a diagram of the magnetic particles 50 adsorb onto Calcium oxalate 110. It means that if the urine sample is displayed with Calcium or Magnesium in urinary crystals, such as calcium oxalate, the urinary crystals can be separated by attaching to the magnetic particles 50 and adding an external magnetic field.

[0073] Otherwise, the present invention provides an urinary crystal detection method. Please refer to FIG. 7 showing a flow chart of the urinary crystal detection method and the steps are as following:

[0074] Step A. An urine sample are provided. In an embodiment, the amount of the urine sample is 1-5 ml from a patient's urine.

[0075] Step B. The urine sample is detected to get a pH value.

[0076] Step B. The urine sample is detected to get a pH value.

[0077] Step C, Processes are executed according to the pH value. If the pH value of urine sample 10 is less than 7, the urine sample is executed acid processes. And the pH value of urine sample 10 is larger than 7, the urine sample is executed alkali processes. In an embodiment, the acid processes comprise a step of adding 5%˜15% acetic acid to the urine sample, and the alkali processes comprise a step of adding 5%˜15% NaOH to the urine sample. After pretreatments, the urine sample Ruins urinary crystals, which is not limited herein.

[0078] Step D. A plurality of magnetic particles are added into the urine sample, the magnetic particles adsorbs urinary crystals of the urine sample.

[0079] Step E. A magnetic field is provided to the urine sample. In an embodiment, the urine sample is provided an external magnet to attract the magnetic particles.

[0080] Step F. The magnetic particles attracted are collect by the magnetic field.

[0081] Step G. The types of the urinary crystals are detected, and comparing the types of urinary crystals with a component information from a database to receive a result of the urinary calculus.

[0082] In an embodiment, a Raman signal of the urinary crystals is detected to get an analyzing result used to determine the type of the urinary crystals, and the peak characteristics of the Raman signals are compared with a component information from a database, wherein the component information is a plurality of reference peak characteristics from different urinary crystals related to a plurality of urinary calculus. It can receive a result of the urinary calculus which the reference peak characteristics is similar as the Raman signal.

[0083] The urinary crystal detection method can be executed step by step (above Step A to Step G orderly), and also be executed from step A and then directly performing step D. Each step can be adjusted according to different cases, which is not limited herein.

[0084] As above embodiment, please refer to FIG. 8A and FIG. 8B showing an urinary calculus patient's analyzing results from the urinary crystal detection system and method of the invention. FIG. 8A is a diagram of urinary crystals under a microscope, and the status of the urinary crystals is calcium oxalate dihydrate (COD). FIG. 8B is a diagram of Raman signals from COD. In FIG. 8B, it can show that the peak characteristics of Raman signals in COD are 910 (cm.sup.−1) and 1477 (cm.sup.−1).

[0085] In another embodiment of the present invention, please refer to FIG. 9A and FIG. 9B showing another urinary calculus patient's analyzing results from the urinary crystal detection system and method of the invention. FIG. 9A is a diagram of urinary crystals under the microscope, and the status of the urinary crystals is hydroxyapatite (HAP). FIG. 9B is is a diagram of Raman signals of the HAP. In FIG. 9B, it can show that the peak characteristic of Raman signals in HAP is 960 (cm.sup.−1).

[0086] The urinary crystal detection system and method of the invention can be used to detect all kinds of urinary crystals, and construct a database via the analyzing results and further to be the reference for comparison afterwards. For example, in an embodiment of the invention, a database is constructed by Raman signals from all kinds of urinary crystals, as shown in table 1, but which is not limited herein.

TABLE-US-00001 TABLE 1 the database of Raman signals. peak characteristics of Urinary crystal/calculus Raman signals (cm.sup.−1) COM (calcium oxalate monohydrate) 895 {grave over ( )} 1462 COD (calcium oxalate dihydrate) 910 {grave over ( )} 1477 DCPD (dicalcium phosphate dihydrate) 986 HAP(hydroxyapatite) 960 Uric acid 623 {grave over ( )} 996 {grave over ( )} 1036 Struvite (magnesium ammonium phosphate) 563 {grave over ( )} 944 Cystine 498

[0087] In another embodiment, an auto-fluorescence signal of the urinary crystals is detected to get an analyzing result by a fluorescence spectroscopy unit. The auto-fluorescence happening is because of organic matter, protein or other substances doped in the formation of the crystallization process, and the crystalline surface also may adsorb protein. Auto-fluorescence signal may be excited from different excitation bands, from UV to near infrared light. It is presumed that the urinary crystals may aggregate to form the urinary stones. In the invention, we use different wavelengths of light to excite crystallization to produce different wavelengths of auto-fluorescence, and determine the composition of crystallization (or crystalline surface) organic matter. It is presumed whether the urinary crystals have the risk to aggregate stones.

[0088] In sum, the urinary crystal detection system and method of the invention can efficiently collect and detect the urinary crystals in the urine. The urine of an urinary calculus patient generally has urinary crystals. Custom made magnetic nanoparticles and an external magnetic field can be used to collect and separate urinary crystals/micro urine stones from other substances in urine. The types of urinary crystals can be determined by the Raman signals. Early detection can be achieved for urinary calculus patients by measuring their urinary crystal. For emergency cases, if the patient has urinary calculus symptoms, this invention can also provide real-time diagnostic efficacy. The invention has the advantages of assisting clinical examination to determine the type of the urinary crystals that may be hard to determine. It is more accurate, more specific, and can assist clinicians to provide individual treatment for the patients according to different types of urinary crystals. It is also convenient to keep tracking and monitoring the patients after the urine stone surgery and to reduce the relapse rate.

[0089] Although the present invention has been described in terms of specific exemplary embodiments and examples, it will be appreciated that the embodiments disclosed herein are for illustrative purposes only and various modifications and alterations might be made by those skilled in the art without departing from the spirit and scope of the invention as set forth in the following claims.